scholarly journals Good Cell Culture Practice for stem cells and stem-cell-derived models

ALTEX ◽  
2016 ◽  
Author(s):  
David Pamies
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Stem Cell ◽  
Good Cell

scholarly journals Feasibility of mesenchymal stem cell culture expansion for a phase I clinical trial in multiple sclerosis

2018 ◽  
Vol 4 (1) ◽  
pp. 205521731876528 ◽  
Author(s):  
Sarah M Planchon ◽  
Karen T Lingas ◽  
Jane Reese Koç ◽  
Brittney M Hooper ◽  
Basabi Maitra ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Stem Cells ◽  
Cell Culture ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Mesenchymal Stem Cell ◽  
Phase I ◽  
Mesenchymal Stem Cell Culture ◽  
Stem Cell Culture ◽  
Multiple Sclerosis Patients

Background Multiple sclerosis is an inflammatory, neurodegenerative disease of the central nervous system for which therapeutic mesenchymal stem cell transplantation is under study. Published experience of culture-expanding multiple sclerosis patients’ mesenchymal stem cells for clinical trials is limited. Objective To determine the feasibility of culture-expanding multiple sclerosis patients’ mesenchymal stem cells for clinical use. Methods In a phase I trial, autologous, bone marrow-derived mesenchymal stem cells were isolated from 25 trial participants with multiple sclerosis and eight matched controls, and culture-expanded to a target single dose of 1–2 × 106 cells/kg. Viability, cell product identity and sterility were assessed prior to infusion. Cytogenetic stability was assessed by single nucleotide polymorphism analysis of mesenchymal stem cells from 18 multiple sclerosis patients and five controls. Results One patient failed screening. Mesenchymal stem cell culture expansion was successful for 24 of 25 multiple sclerosis patients and six of eight controls. The target dose was achieved in 16–62 days, requiring two to three cell passages. Growth rate and culture success did not correlate with demographic or multiple sclerosis disease characteristics. Cytogenetic studies identified changes on one chromosome of one control (4.3%) after extended time in culture. Conclusion Culture expansion of mesenchymal stem cells from multiple sclerosis patients as donors is feasible. However, culture time should be minimized for cell products designated for therapeutic administration.

scholarly journals Stem cell culture on polyvinyl alcohol hydrogels having different elasticity and immobilized with ECM-derived oligopeptides

2017 ◽  
Vol 37 (7) ◽  
pp. 647-660 ◽  
Author(s):  
Saradaprasan Muduli ◽  
Li-Hua Chen ◽  
Meng-Pei Li ◽  
Zhao-wen Heish ◽  
Cheng-Hui Liu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Extracellular Matrix ◽  
Stem Cell ◽  
Cell Fate ◽  
Es Cells ◽  
Embryonic Stem ◽  
Poly Vinyl Alcohol ◽  
Hematopoietic Stem ◽  
Stem Cell Culture

Abstract The physical characteristics of cell culture materials, such as their elasticity, affect stem cell fate with respect to cell proliferation and differentiation. We systematically investigated the morphologies and characteristics of several stem cell types, including human amniotic-derived stem cells, human hematopoietic stem cells, human induced pluripotent stem (iPS) cells, and embryonic stem (ES) cells on poly(vinyl alcohol) (PVA) hydrogels immobilized with and without extracellular matrix-derived oligopeptide. Human ES cells did not adhere well to soft PVA hydrogels immobilized with oligovitronectin, whereas they did adhere well to PVA hydrogel dishes with elasticities greater than 15 kPa. These results indicate that biomaterials such as PVA hydrogels should be designed to possess minimum elasticity to facilitate human ES cell attachment. PVA hydrogels immobilized with and without extracellular matrix-derived oligopeptides are excellent candidates of cell culture biomaterials for investigations into how cell culture biomaterial elasticity affects stem cell culture and differentiation.

scholarly journals Growing, Tracking, and Directing Bone Marrow Derived Stem Cells From Two-Dimensional and Three-Dimensional Cell Culture Microenvironments

2019 ◽  
Author(s):  
Tiffany Miller
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Bone Marrow ◽  
Stem Cell ◽  
Three Dimensional ◽  
Inflammatory Biomarkers ◽  
Two Dimensional ◽  
Cell Culturing ◽  
Dimensional Cell ◽  
Stimulation Of

<p>Bone marrow derived stem cells express biomarkers capable of facilitating adhesion to the cell culturing microenvironment, thereby, influencing their proliferation, migration, and differentiation. In particular, biological biomarkers of mesenchymal stem cells include, but are not limited to, CD14-, CD19-, CD34-, CD45-, CD29, CD44, CD73+, CD90+, CD105+, CD106, CD166, Stro-1, and HLADR. The relationship between the stem cell biology and the materials and methods forming a cell culturing microenvironment serves as a critical aspect in the successful adhesion and growth within two-dimensional cell culture microenvironments such as polystyrene, laminin, fibronectin, or poly-L-lysine and within three-dimensional cell culture microenvironments such as hydrogel, ceramic, collagen, polymer based nanofibers, agitation, forced floating, and hang drop systems. Further, electrical stimulation of the stem cells may be implemented during the cell culturing process to measure stem cell growth and to determine stem cell viability. In addition, electrical stimulation of implanted stem cells may facilitate tracking by measuring stem cell migration distance and travel area. Although many biochemical and inflammatory biomarkers are expressed based on severity in stroke including, but not limited to, Interluken-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and glutamate (Glu), current methodologies of stem cell directing lack localization and biological effector specificity. Biological effector bound magnetic particle stem cells may serve as a potential treatment method in ischemic stroke. In particular, a stem cell biomarker may be configured to communicate with inflammatory biomarkers, thus, more efficiently delivering the stem cells to site specific areas having the most severely affected <i>in-vivo</i> biochemical microenvironments.</p>

289 STEM CELL CULTURE AND DIFFERENTIATION IN MICROFLUIDIC DEVICES

2013 ◽  
Vol 25 (1) ◽  
pp. 292
Author(s):  
E. Khetan ◽  
A. J. Maki ◽  
M. B. Wheeler
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Stem Cell ◽  
Toluidine Blue ◽  
Microfluidic Devices ◽  
Great Promise ◽  
Small Scale ◽  
Microfluidic Channels ◽  
Alizarin Red ◽  
Oil Red O

Advancements in micro and nanotechnology have allowed scientists a powerful platform to study biological systems. Microfluidics is one area of advancement with great promise. Microfluidics deals with the behaviour, specific control, and manipulation of microliter and nanoliter volumes of fluid. The small-scale design of these microfluidic devices permits laminar flow, characterised as parallel streams flowing without disruption between currents. With the introduction of micro-technology and microfluidic platforms for cell culture, stem cell research can be put into a new context. Inside microfluidics, microenvironments can be more precisely controlled and they provide a more in vivo-like environment for the cells to grow and hence can serve as a better way of culturing the cells. In the current study, we examined the influence of microfluidic devices on the development of stem cells. Adipose-derived stem cells (ADSC) were isolated from pigs and seeded in a microfluidic device to differentiate toward adipogenic, osteogenic, and chondrogenic lineages using specific differentiation-promoting media (Monaco et al. 2009 Open Tissue Eng. Regen. Med. J. 2, 20–33). Five thousand cells were seeded per channel at a density of 5 000 000 cells mL–1. The microchannel dimensions were 5 mm long, 1 mm wid, and 200 µm deep. Cells were maintained for 14 days and then stained with respective staining dyes: Oil Red O for adipogenesis, Alizarin Red for osteogenesis, and Toluidine Blue for chondrogenesis. Cells differentiated towards adipogenic lineage contained small lipid droplets, which stained red with Oil Red O stain; during osteogenic differentiation, the cells formed large nodules and stained positive for the presence of calcium; and the chondriogenic differentiating cells showed the presence of proteoglycans (blue) when stained with Toluidine Blue. We seeded ADSC in 5 channels for each differentiation lineage, and all channels gave positive staining results. We conclude that microfluidic channels support proliferation and differentiation of ADSC. This system uses small amounts of culture medium, experiments with different culture compositions can be efficiently performed, and culture manipulations can be automated using fluid-handling robotics. Because microfluidics can deal with small number of cells, the characteristics of cellular structure and function and the microenvironment of the stem cells can be understood in a more precise manner. The miniaturization of cell culture platforms allows the observation of cellular behaviour at the scale found in living systems.

E-Cadherin – Fc Chimeric Protein-Based Biomaterial: Breaking the Barriers in Stem Cell Technology and Regenerative Medicine

2013 ◽  
Vol 810 ◽  
pp. 41-76 ◽  
Author(s):  
Kakon Nag ◽  
Toshihiro Akaike
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Stem Cell ◽  
Regenerative Medicine ◽  
Chimeric Protein ◽  
Biochemical Properties ◽  
Target Cells ◽  
Chimeric Proteins ◽  
Stem Cell Technology ◽  
Cell Technology

Chimeric proteins have been used for years for various purposes ranging from biomaterials to candidate drug molecules, and from bench to bulk. Regenerative medicine needs various kinds of proteins for providing essential factors for maintaining starting cells, like induced pluripotent stem cells (iPSC), and renewal, proliferation, targeted differentiation of these cells, and as extracellular matrix for the experimental cells. However, there are several challenges associated with making functional chimeric proteins for effective application as biomaterial in this field. Fc-chimeric protein technology could be an effective solution to overcome many of them. These tailored proteins are recently becoming superior choice of biomaterials in stem cell technology and regenerative medicine due to their specific advantageous biophysical and biochemical properties over other chimeric forms of same proteins. Recent advances in recombinant protein-related science and technology also expedited the popularity of this kind of engineered protein. Over the last decade our lab has been pioneering this field, and we and others have been successfully applied Fc-chimeric proteins to overcome many critical issues in stem cell technologies targeting regenerative medicine and tissue engineering. Fc-chimeric protein-based biomaterials, specifically, E-cad-Fc have been preferentially applied for coating of cell culture plates for establishing xenogeneic-agent free monolayer stem cell culture and their maintenance, enhanced directed differentiation of stem cells to specific lineages, and non-enzymatic on-site one-step purification of target cells. Here the technology, recent discoveries, and future direction related with the E-cad-Fc-chimeric protein in connection with regenerative medicine are described.

scholarly journals Growing, Tracking, and Directing Bone Marrow Derived Stem Cells From Two-Dimensional and Three-Dimensional Cell Culture Microenvironments

2019 ◽  
Author(s):  
Tiffany Miller
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Bone Marrow ◽  
Stem Cell ◽  
Three Dimensional ◽  
Inflammatory Biomarkers ◽  
Two Dimensional ◽  
Cell Culturing ◽  
Dimensional Cell ◽  
Stimulation Of

<p>Bone marrow derived stem cells express biomarkers capable of facilitating adhesion to the cell culturing microenvironment, thereby, influencing their proliferation, migration, and differentiation. In particular, biological biomarkers of mesenchymal stem cells include, but are not limited to, CD14-, CD19-, CD34-, CD45-, CD29, CD44, CD73+, CD90+, CD105+, CD106, CD166, Stro-1, and HLADR. The relationship between the stem cell biology and the materials and methods forming a cell culturing microenvironment serves as a critical aspect in the successful adhesion and growth within two-dimensional cell culture microenvironments such as polystyrene, laminin, fibronectin, or poly-L-lysine and within three-dimensional cell culture microenvironments such as hydrogel, ceramic, collagen, polymer based nanofibers, agitation, forced floating, and hang drop systems. Further, electrical stimulation of the stem cells may be implemented during the cell culturing process to measure stem cell growth and to determine stem cell viability. In addition, electrical stimulation of implanted stem cells may facilitate tracking by measuring stem cell migration distance and travel area. Although many biochemical and inflammatory biomarkers are expressed based on severity in stroke including, but not limited to, Interluken-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and glutamate (Glu), current methodologies of stem cell directing lack localization and biological effector specificity. Biological effector bound magnetic particle stem cells may serve as a potential treatment method in ischemic stroke. In particular, a stem cell biomarker may be configured to communicate with inflammatory biomarkers, thus, more efficiently delivering the stem cells to site specific areas having the most severely affected <i>in-vivo</i> biochemical microenvironments.</p>

P04.19 A combined use of a G-quadruplex oligonucleotide (GQ) and neuro-inducing small molecules inhibit glioblastoma cell proliferation

2021 ◽  
Vol 23 (Supplement_2) ◽  
pp. ii22-ii22
Author(s):  
V Kolesnikova ◽  
N Samoylenkova ◽  
S Drozd ◽  
A Revishchin ◽  
D Y Usachev ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Cell Proliferation ◽  
Stem Cell ◽  
Cell Cultures ◽  
Mtt Assay ◽  
Proliferative Activity ◽  
Recombinant Dna ◽  
Glioblastoma Cell ◽  
G Quadruplex

Abstract BACKGROUND According to one of the theories, gliomas can occur as a result of dysregulation of stem cell division in the subventricular region of the brain. The CD133 membrane marker is a characteristic of both normal and tumor neural stem cells therefore it can be used to isolate a stem cell population from tumor tissue. Tumor cells actively proliferate which suggests that their possible differentiation may be achieved by inhibiting of their division as these two processes are mutually exclusive. For this purpose, G-quadruplex oligonucleotides together with neural-inducers such as a brain-derived neurotrophic factor (BDNF) may be used. MATERIAL AND METHODS Five cell cultures obtained from human glioblastoma tissues were analyzed for expression of CD133 using RT-qPCR. From cell culture with the highest level of CD133 using immunomagnetic separation CD133+ and CD133- cultures were received. CD133fr/peGFP-c1 recombinant DNA consisted of a CD133 second extracellular loop fragment and a peGFP-c1 vector was constructed to determine the localization of prominin-1, that is known as CD133 when found on cell membrane, using confocal microscopy. On chosen cell cultures an oligonucleotide bi-(AID-1-T) and its combination with BDNF were tested. The mechanism of GQ’s action is cytostatic and its non-toxicity properties were proved by flow cytometry. For evaluating the proliferative activity of cells MTT assay was performed on 10th and 20th days after exposure to the factors. RESULTS Cell culture G01 was chosen for further research as it had the highest level of the CD133. Colocalization of CD133 and GFP demonstrated a membrane localization of CD133 in cells with high expression level of this marker. MTT assay on 10th day after exposure to bi-(AID-1-T) as well as its combination with BDNF on cell culture G01 CD133- showed total inhibition of cell proliferation. The same combinations tested on G01 CD133+ cell culture demonstrated no difference in proliferative activity. After 20 days after exposure to bi-(AID-1-T) and combination of bi-(AID-1-T) with BDNF the significant decrease of G01 CD133+ cells’ proliferation was observed. When tested on whole glioblastoma cell culture G01 these combinations also showed significant inhibition of cell proliferation. CONCLUSION We showed that glioblastoma cells upon transfection with recombinant DNA, that contains a fragment of CD133, mainly have a membrane localization of this marker. It was observed that CD133+ cells are more stable to external influence that can be a proof of the fact that CD133 is charactered for glioblastoma stem cells. We tested the effect of an GQ bi-(AID-1-T) and its combination with BDNF and showed that BDNF is necessary for blocking proliferation of glioblastoma cells. Altogether, the results may be used for further research as it reveals a potential treatment for patients with glioblastoma. Grant №075-15-2020-809 (13.1902.21.0030).

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